Abstract
BackgroundLung cancer is one of the most lethal malignancies worldwide, but useful biomarkers of lung cancer are still insufficient. The aim of this study is to identify some membrane-bound protein(s) associated with migration and invasion in human non-small cell lung cancer (NSCLC) cells.MethodsWe classified four NSCLC cell lines into high and low migration/invasion groups by Transwell and Matrigel assays. Using two-dimensional gel electrophoresis and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS), we identified 10 membrane-associated proteins being significantly overexpressed in the high migration/invasion group. The expression of the target protein in the four NSCLC cell lines was then confirmed by reverse transcription polymerase chain reaction (RT-PCR), western blot and immunostaining. RNA interference technique was applied to observe the influence of the target protein on migration and invasion. Gelatin zymography was also performed to evaluate the activities of matrix metalloproteinase (MMP)-2 and MMP-9. Expression condition of the target protein on surgical specimens was further examined by immunohistochemical staining and the clinicopathologic data were analyzed.ResultsWe identified a mitochondria-bound protein cytochrome c oxidase subunit Va (COX Va) because of its abundant presence found exclusively in tumorous areas. We also demonstrated that migration and invasion of NSCLC cells decreased substantially after knocking down COX Va by siRNA. Meanwhile, we found a positive correlation between COX Va expression, Bcl-2 expression and activities of MMP-2 and MMP-9 in NSCLC cells. Immunohistochemical staining of surgically resected lung adenocarcinomas in 250 consecutive patients revealed that strong COX Va expression was found in 54.8% (137/250) of patients and correlated positively with the status of lymph node metastasis (P = 0.032). Furthermore, strong COX Va expression was associated with the presence of distant metastasis (P = 0.033).ConclusionsOur current study showed that COX Va may play a role in migration and invasion of NSCLC cells and can be used as a biomarker to predict aggressiveness of NSCLC.
Highlights
Lung cancer is one of the most lethal malignancies worldwide, but useful biomarkers of lung cancer are still insufficient
Characterization of molecular and protein expression of c oxidase subunit Va (COX Va) in non-small cell lung cancer (NSCLC) cell lines by reverse transcription polymerase chain reaction (RT-PCR), western blot and immunocytochemical staining To evaluate whether this newly identified protein c oxidase (COX) Va was overexpressed in situ in the cells with high migration/
matrix metalloproteinase (MMP) assay To look into the factors that were possibly linked to COX Va expression, we examined the expression of matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase9 (MMP-9) in these four NSCLC cells
Summary
Lung cancer is one of the most lethal malignancies worldwide, but useful biomarkers of lung cancer are still insufficient. About 40-50% patients of NSCLC present with stage IV disease [5], and given that complete surgical resection may provide a chance of cure in patients with early-stage tumors, the reported recurrence rate in the patients with completely resected stage I NSCLC was nearly 30% [6,7,8]. Among these patients with tumor recurrence, more than 70% of them have distant metastasis [6,8]. Molecular factors that are related to invasion are still insufficient, and identification of such factors with elucidation of their molecular mechanism will provide insight into cancer biology and potentially provide new therapeutic targets for NSCLC patients
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